PROJECT SUMMARY/ABSTRACT A major problem in the field of eukaryotic infectious diseases remains unsolved: why are some parasites pathogenic but other closely related ones relatively benign? Here we propose to address this problem by studying malaria, a diverse suite of pathogens where some cause severe morbidity yet others can be asymptomatic. By studying the gene expression of malaria parasites that infect birds, we will elucidate the evolution of pathogenicity and identify genes that cause morbidity. We will test the hypothesis that certain genes are differentially expressed between virulent and benign pathogens. Using a bioinformatics approach, we will compare the transcriptomes of avian pathogens and identify whether different genes are expressed, and/or if they are expressed at different levels between the virulent and non-virulent Plasmodium species. By selectively mutating avian malaria genes, we will verify the alleles responsible for pathogenesis. The CRISPR/Cas9 approach has been used recently to disrupt genes in Plasmodium falciparum. This will be the first use of this approach in a non-mammalian malaria system. The proposed research is timely and significant in that it will advance the field of malaria research by providing an important new model system to identify factors contributing to malaria pathogenicity. This work is innovative because birds provide the only model system where two highly related malaria parasites cause such fundamentally different outcomes to both laboratory and naturally occurring wild hosts. The long-term and cross-disciplinary goal of this research agenda is to elucidate the evolution of pathogenicity and identify conserved virulence mechanisms that may be overlooked using mammalian model systems. The aims of the proposal are to: 1. Sequence the complete transcriptomes of select avian malaria parasites. 2. Characterize differences in gene expression between virulent and non-virulent parasite strains. 3. Use a CRISPR/Cas9 approach to disrupt virulence genes and confirm effects through the infection of laboratory birds. The project will support the education of future scientists through an integrative approach to research and education. It will train students in cutting edge technologies associated with infectious disease research, and provide valuable experiences for graduate and undergraduate students at an urban university serving large numbers of underrepresented minority students. The developmental objectives of the PI will be met by working with outstanding collaborators, which will further his training in the genomics of malaria.